Impairment of myocardial angiogenesis and coronary collateral growth may contribute to high mortality in diabetic myocardial infarction. Our long-term goal is to define the molecular mechanism(s) responsible for abnormal vascular maturation and impairment of angiogenesis in the diabetic hearts. This revised proposal will investigate a possible disruption in the angiopoietins (Ang)/Tie-2 and apelin pathway in abnormal diabetes- associated vascular maturation and capillary regression. Our laboratory has shown a sustained increase in angiopoietin-2 (Ang-2) and prolyl hydroxylase-2 (PHD2) expression, and reduced Ang-1/Tie-2 and HIF- 11/apelin expression in diabetic mice. Our previous demonstration of impaired myocardial vessel maturation in diabetic mice; implicate that disruption of angiopoietins/Tie-2 system in favor of Ang-2, which leading to immature vessel formation and capillary regression, might be a novel mechanism responsible for impaired angiogenesis in diabetic hearts. Our overall hypothesis is that diabetes disrupts Ang-1/Tie-2 and apelin pathway by a mechanism involving Ang-2 and PHD2 activation; and these abnormalities lead to abnormal vascular maturation and capillary regression in diabetic hearts. Specific Aim 1 will define the mechanism(s) by which hyperglycemia interferes with vascular maturation and capillary regression with a focus on the role of Ang-2 in the disruption of Ang-1/Tie-2 and apelin pathway. Using heart microvascular endothelial cells (EC), co-cultured EC-SMC spheroids and mouse aortic explants isolated from wild type (WT) or diabetic db/db mice, we will determine whether: (i) high glucose-induced excess of Ang-2 disrupts Ang-1/Tie-2 signaling and attenuates Ang-1-induced apelin expression; and (ii) interactions between Ang-2 and apelin are critical for the regulation of angiogenesis and vascular regression under high glucose conditions. In specific aim 2, we will determine the role of Ang-2 and PHD2 activation in diabetes-associated disruption of vascular maturation and angiogenesis and promotion of vessel regression in an in vivo model of myocardial ischemia. Using Ang-2 deficient and PHD2 conditional knockout diabetic mice models, we will determine whether deficiency of Ang-2 or endothelial cell deletion of PHD2 rescues impaired apelin expression, normalizes immature neovessels, and improves myocardial angiogenesis. In specific aim 3, we will further determine whether systemic administration of apelin rescues impaired angiogenic signaling, normalizes immature neovessels, and increases myocardial angiogenesis in diabetic hearts. Our studies will provide a framework for the development of a targeted therapeutic reduction in Ang-2 and PHD2 activation to ameliorate or reverse the abnormalities in diabetic vessel maturation and angiogenesis that characterizes the diabetic state.